Early repair of defects in articular cartilage could prevent the development of osteoarthritis. Autologous, cultured chondrocytes have been implanted in human articular cartilage defects with some success early on but with questionable durability due to a failure to develop true hyaline cartilage. Cell expansion in vitro typically leads to the loss of the cartilage phenotype. The long term goal of this project is to develop a better cartilage transplant for repair of defects in articular cartilage. Our plan is to combine two novel culture techniques, each of which have been shown to have promise on their own. These techniques include isolation of chondrons (chondrocytes plus their native pericellular matrix( instead of the traditional approach of using chondrocytes stripped of all extracellular matrix. Matrix interactions have been shown to play a major role in the phenotypic expression of cells in vitro. The second culture technique is to culture the chondrons under a dynamic mechanical load. Mechanical load also affects phenotypic expression and has been shown to promote expression of cartilage specific matrix macromolecules. Our strategy will be to use state-of-the-art differential display techniques as an indicator of retention of the cartilage phenotype. Such a precise indicator will allow fine tuning of the culture process. Our hypothesis is that chondrons cultured with the application of a dynamic mechanical load will produce a more hyaline cartilage-like matrix than will nonloaded chondrocytes. The specific aims of this proposal are: (1) determine the pattern of messenger RNA expression by chondrocytes in normal and osteoarthritic human articular cartilage to identify markers unique to normal cartilage. (2) determine the optimum conditions of dynamic mechanical loading to maintain loading to maintain the normal cartilage phenotype in culture using the unique markers identified in Specific Aim 1 as the gold standard (3) determine the optimum growth factor supplements for promoting growth and matrix production while maintaining the cartilage phenotype in a mechanically loaded culture. Chondrons will be enzymatically isolated from normal human cartilage and cultured under mechanical load. Differential display PCR will be used to identify unique gene sequences through cloning and sequencing of novel genes.